Our broad objective is to understand the means by which Borrelia burgdorferi (Bb) establishes an infection:transmission cycle between the tick vector and mammalian reservoir host, both of which are needed to maintain the spirochete in nature. 1. Outer surface protein variation. The outer membrane of Bb contains several abundant proteins (Osps) that vary in size and expression and are of unknown function. It is likely that the different Osps confer distinct properties on the spirochete that are pertinent to the different environments in which it must survive. Collaborative studies with Dr. Schwan have shown that OspC is detectable on spirochetes in the midgut of infected ticks only after a bloodmeal, and that an increase in temperature is in part responsible for increased OspC synthesis. Dr. Stevenson has documented that at least 5 other proteins are differentially synthesized as a consequence of culture temperature. Dr. Stevenson has analyzed the 5' flanking sequence of the ospC gene from multiple strains to identify conserved promoter elements. Fisher, Stevenson and Tilly have initiated studies using both genetic and biochemical approaches to identify components that alter osp promoter activity. Stevenson, Tilly and Rosa are attempting to transform Bb and inactivate osp genes and other genes of interest by integrating selectable markers. Such knockout mutants are a critical component of future studies to determine the roles of various proteins, including Osps, in the infectious cycle of Bb. 2. Plasmid structure and replication. Understanding the structure and replication of the unusual linear and circular Borrelia genome is of intrinsic interest and practical merit. Dr. Tilly has continued her studies of a Bb HU/IHF homolog. She and Dr. Samuels (RMLMB) have developed a purification scheme and used a gel shift assay to identify protein fractions that bind to a telomeric DNA fragment that contains a putative IHF binding site. Tilly and Rosa have developed a strategy to analyze the site at which telomeric sequences recombine. Rosa and Tilly have extended their analysis of the structure and function of the unique plasmid-encoded Bb IMP dehydrogenase gene homolog (guaB) by comparing it to the closely related, but more typical, homolog in B. hermsii. Previous studies have identified a gene homolog of a peptide binding protein adjacent to guaB on the 26-kB circular plasmid. Dr. Rosa has extended this study by identifying a chromosomal locus homologous to an oligopeptide permease system.